As a part of undergrad lab project, you have to clone Your Favorite Gene into plasmid cloning vector (4740 bp) pUC18. Restriction maps for both YFG and cloning vector are provided.The plasmid has a bacterial origin of replication (ori), an ampicillin resistance gene (AmpR) that degrades ampicillin antibiotic. It also has LacZ gene that encodes the β-gal enzyme, which converts white X-gal substrate into a blue product. pLac is a bacterial promoter.Restriction Enzyme Recognition Site : A slash (/) represents the cut site for each restriction enzyme.Enzyme 15’- G/TCGA C -3’3’- C AGCT/G-5’ Enzyme 25’-C/TCGA G-3’3’G AGCT/ C5’Enzyme 35’- GTC / GAC- 3’3’- CAG / CTG- 5’Enzyme 45’-C AATT /G-3’3’-G/ TTAA C-5’ Enzyme 55’C AATT/G-3’3’G/TTAA C-5’Enzyme 65’G/GATC C-3’3’C CTAG/ G-5’ Enzyme 75’G/GATC C-3’3’C CTAG/ G-5’1a)The table below shows restriction enzyme pairs used by various teams to digest the YFG and clone it in the bacterial plasmid cut with the same enzymes.Complete the table below for each restriction enzyme pair. Restriction enzymes used to cut YFGWill you be able to…Column 1: Clone the digested YFG into the plasmid – YES/NO?Column 2: Amplify the recombinant plasmid in the ampicillin sensitive bacterial cells:YES/NO?Column 3: Express YFG in the bacterial cells: YES/NO/Maybe?1 and 5 1 and 2 2 and 6 3 and 7 The image shows a diagram of the pUC18 plasmid, which is a circular DNA molecule commonly used in cloning. This plasmid consists of 2686 base pairs (bps). Key features are highlighted in the diagram:1. **Amp**: This is the gene that provides ampicillin resistance, shown as a green arrow, which is critical for selecting bacteria that have been successfully transformed with this plasmid.2. **pMB1 ori**: The origin of replication, depicted in light blue, is where DNA replication begins, allowing the plasmid to be copied within a bacterial cell.3. **lacZ**: Shown as a dark blue arrow, this gene is part of the lac operon and codes for beta-galactosidase, an enzyme that can be used to identify successful cloning via blue-white screening.4. **MCS (Multiple Cloning Site)**: This is a short sequence containing multiple unique restriction enzyme sites where foreign DNA can be inserted. It is positioned near the lacZ gene, as indicated by the location of enzyme sites.5. **LacR binding site**: Located near the MCS, this site interacts with the Lac repressor, regulating expression from the lacZ gene.6. **Plac**: The promoter for the lacZ gene is marked in red, controlling the initiation of transcription.7. The box titled "Enzymes sites (in order)" lists enzyme sites numerically from 1 to 7, corresponding to the specific sites within the MCS, although these specific enzymes are not named in the diagram.This diagram is critical for understanding the functions and applications of the pUC18 plasmid in genetic cloning experiments.

Answered: Image /qna-images/answer/1dfd2526-752b-4642-8857-87902dc5918f.jpg

Answered: The table below shows restriction…

Biochemistry

9th Edition

ISBN:9781319114671

Author:Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.

Publisher:Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.

Chapter1: Biochemistry: An Evolving Science

Section: Chapter Questions

Problem 1P

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Question

As a part of undergrad lab project, you have to clone Your Favorite Gene into plasmid cloning vector (4740 bp) pUC18. Restriction maps for both YFG and cloning vector are provided.

The plasmid has a bacterial origin of replication (ori), an ampicillin resistance gene (AmpR) that degrades ampicillin antibiotic. It also has LacZ gene that encodes the β-gal enzyme, which converts white X-gal substrate into a blue product. pLac is a bacterial promoter.

Restriction Enzyme Recognition Site : A slash (/) represents the cut site for each restriction enzyme.

Enzyme 1	5’- G/TCGA C -3’ 3’- C AGCT/G-5’
Enzyme 2	5’-C/TCGA G-3’ 3’G AGCT/ C5’
Enzyme 3	5’- GTC / GAC- 3’ 3’- CAG / CTG- 5’
Enzyme 4	5’-C AATT /G-3’ 3’-G/ TTAA C-5’
Enzyme 5	5’C AATT/G-3’ 3’G/TTAA C-5’
Enzyme 6	5’G/GATC C-3’ 3’C CTAG/ G-5’
Enzyme 7	5’G/GATC C-3’ 3’C CTAG/ G-5’

1a)

The table below shows restriction enzyme pairs used by various teams to digest the YFG and clone it in the bacterial plasmid cut with the same enzymes.

Complete the table below for each restriction enzyme pair.

Restriction enzymes used to cut YFG	Will you be able to…
Column 1: Clone the digested YFG into the plasmid – YES/NO?	Column 2: Amplify the recombinant plasmid in the ampicillin sensitive bacterial cells: YES/NO?	Column 3: Express YFG in the bacterial cells: YES/NO/Maybe?
1 and 5
1 and 2
2 and 6
3 and 7